白色念珠菌分泌型天冬氨酸蛋白酶的纯化与性质

利用硫酸铵分级沉淀和DEAE-Sephacel离子交换层析从白色念珠菌WD27培养液中提纯得到一种蛋白酶,纯化倍数为25.4,收率为5.2％。其活性可被抑胃肽特异抑制,初步鉴定该酶为天冬氨酸蛋白酶。该酶在酸性范围内有水解蛋白活性,最适作用pH为4.0,最适温度为37℃。酶在pH5.0～6.0、45℃以下较稳定,该酶具有较广的底物作用范围,对牛血红蛋白最敏感。酶作用于牛血红蛋白的米氏常数Km为0.814mmol/L。     

Clara cell protein (CC16) in serum and bronchoalveolar lavage fluid of subjects exposed to asbestos

The Clara cell protein (CC16) is a small and readily diffusible protein of 16kDa secreted by bronchiolar Clara cells in the distal airspaces. These epithelial cells are altered in several pulmonary pathological processes induced by various lung toxicants. In the search for a new biomarker of asbestos-induced lung impairment, we used a sensitive immunoassay to determine the levels of CC16 in bronchoalveolar fluid (BALF) and serum of subjects exposed to asbestos compared with a group of healthy controls. In the BALF of asbestos-exposed subjects there was an insignificant trend towards CC16 elevation compared with controls, with a (mean ±SD of 0.81 ±0.65mg l^-1 for asbestos-exposed subjects (n = 23) versus 0.39 ±0.19mg l^-1 for controls (n = 11) (p = 0.09). In serum, CC16 concentration was significantly increased among asbestos-exposed subjects, with values of 27.2 ±24.0 µg l^-1 for asbestos-exposed subjects (n = 34) versus 16.1 ±7.6 µg l^-1 for controls (n = 34) (p = 0.01). Regarding the effects of smoking, there were significant differences between generally lower CC16 levels in serum and BALF (p = 0.05 and 0.001, respectively) of smokers compared with the higher levels in non-smokers. Serum CC16 levels positively correlated with those in BALF, which is consistent with a diffusional transfer of CC16 from the bronchoalveolar space into the serum. No association, however, emerged between the levels of CC16 in serum or BALF and either the duration of asbestos exposure or the severity of the lung impairment as assessed by chest X-ray. These findings suggest that exposure to asbestos elicits early changes in the local and, importantly, also the systemic levels of CC16. This pneumoprotein therefore appears as a promising non-invasive biomarker of asbestos-induced lung injury and occupational disease in both smoking and non-smoking exposed subjects.     

Alterations in pancreatic islet phosphate content during secretory stimulation with glucose

Isolated rat pancreatic islets were perifused and analyzed for phosphate content immediately following the transient increase in the efflux of orthophosphate which occurs when insulin secretion is stimulated by glucose. In some instances, islets were perifused directly following isolation to minimize preparative delay; in others, islets were prelabeled during incubation with [³²P]orthophosphate for 90 min prior to perifusion. In both experimental situations, total islet phosphate content declined 40–50% following exposure to stimulating concentrations of glucose and initiation of enhanced insulin release. In the experiments with prelabeled islets, tissue content of [³²P]orthophosphate fell to a similar extent so that the specific radioactivity of islet orthophosphate was unaffected. Inhibition of heightened insulin release with Ni²⁺ did not modify the decrements in total or radioactive tissue orthophosphate, thus indicating that these responses to islet stimulation reflect events which are proximal to activated exocytosis. Simultaneous analyses for tissue ATP and ADP demonstrated that the efflux in orthophosphate and reduction in tissue orthophosphate content were not mediated via net changes in islet adenine nucleotides. The observations represent the first documentation that a net reduction of tissue inorganic phosphate is one of the early components of stimulus-secretion coupling in isolated pancreatic islets.     

Morphologische,histochemische und experimentelle Untersuchungen an den parafollikulären Zellen der Schilddrüse

Zusammenfassung Die parafollikulären Zellen der Rattenschilddrüse zeigen eine vom jeweiligen Funktionszustand abhängige Feinstruktur: 1. Zellen mit zahlreichen Granula, einem ausgeprägten Golgi-Apparat, gering entwickeltem granuliertem endoplasmatischem Retikulum und manchmal einigen dichten Körpern mit myelinähnlichen Figuren. 2. Zellen mit wenigen Granula und einem stark entwickelten endoplasmatischen Retikulum mit erweiterten Zisternen; diese Zellen können das Lumen des Follikels erreichen. 3. Einige degranulierte Zellen. — In den Schilddrüsen-Follikeln des Hundes konnten wir nur die ersten beiden Zellformen, aber keine degranulierten parafollikulären Zellen beobachten.Nach Ca⁺⁺-Injektion findet man als Zeichen der Funktionsabhängigkeit der Feinstruktur eine Zunahme der Zellen mit stark entwickeltem endoplasmatischem Retikulum und nur geringer Granulation.Die dichten Körper mit myelinähnlichen Figuren zeigen saure Phosphataseaktivität. Es handelt sich deshalb wahrscheinlich um Restkörper, die aus Autolysosomen entstanden sind. Trotzdem zeigt sich nach Zufuhr von Ca⁺⁺ und anschließender EDTA-Gabe keine eindeutige Zunahme der Lysosomenzahl.Zwischen follikulären und parafollikulären Zellen sind Axonanschnitte zu finden.

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Morphological, histochemical and experimental studies on the parafollicular cells of the thyroid

Summary The parafollicular cells of the thyroid of the rat show different fine structures most likely in relation with different functional states: 1. Some cells contain numerous secretory granules, a well developed Golgi complex, a moderately developed rough endoplasmic reticulum and some dense bodies containing myelin figures. 2. Other parafollicular cells have few granules and a strongly developed rough endoplasmic reticulum with enlarged cisternes. They sometimes reach the lumen of the follicle. 3. Finally, a few parafollicular cells appear degranulated.—In dogs the degranulated parafollicular cells could not be observed.Following administration of Ca⁺⁺ ions there is an increase of cells with strongly developed endoplasmic reticulum and only few granules.The dense bodies with myelin figures show acid phosphatase activity. Most likely they are residual bodies derived from autolysosomes. However, EDTA after stimulation of the cells by Ca⁺⁺ does not significantly increase the number of parafollicular cells containing autolysosomes.Axons can be found between follicular and parafollicular cells.

     

Interaction of antimycin with cytochrome b-561: A study in secretory granules and in plasma membrane isolated from chromaffin cells of bovine adrenal medulla

Cytochrome b-561 in chromaffin granules interacts with antimycin and its -peak shifts 1 nm towards red. When chromaffin granules were treated with Triton X-100 antimycin no effect was observed. Cytochrome b-561 is located in the plasma membrane isolated from the chromaffin cells. The plasma membrane b-561 does not seem to interact with antimycin. A number of NADH or NADPH (acceptor) oxidoreductase activity has been observed in isolated plasma membrane providing clues to the origin of plasma membrane dehydrogenase. The possible role of cytochrome b561 in secretory granules other than its accredited energy conserving electron transport property is projected.     

The production of nematode-immobilizing secretory cells by Climacodon septentrionalis

The ability of Climacodon septentrionalis to immobilize and kill a mycophagous nematode (Aphelenchoides sp.) in vitro is described for the first time. Two isolates produced droplets (20–45 μm in diameter) that formed at the apices of tall, stalked, and branching secretory cells (700–1,500 μm tall). On 2% modified malt extract agar, nematodes became enveloped in the droplets, which restricted their ability to move and resulted in complete immobilization and death within several hours of contact. The rate of decomposition of the nematodes varied considerably, with most individuals persisting for weeks whereas others were degraded within several days and appeared to be colonized by dense hyphal growth. This study provides the first documentation of a non-agaricoid fungus producing secretory cells that are able to immobilize nematodes.     

The comparative morphology of epidermal glands in Pentatomoidea (Heteroptera)

The Heteroptera show a diversity of glands associated with the epidermis. They have multiple roles including the production of noxious scents. Here, we examine the cellular arrangement and cytoskeletal components of the scent glands of pentatomoid Heteroptera in three families, Pentatomidae (stink bugs), Tessaratomidae, and Scutelleridae (shield-backed bugs or jewel bugs). The glands are; (1) the dorsal abdominal glands, (2) the tubular glands of the composite metathoracic gland, and (3) the accessory gland component of the composite metathoracic gland. The dorsal abdominal glands are at their largest in nymphs and decrease in size in adults. The metathoracic gland is an adult-specific gland unit with a reservoir and multiple types of gland cells. The accessory gland is composed of many unicellular glands concentrated in a sinuous line across the reservoir wall. The lateral tubular gland is composed of two-cell units. The dorsal abdominal glands of nymphs are composed of three-cell units with a prominent cuticular component derived from the saccule cell sitting between the duct and receiving canal. The cuticular components that channel secretion from the microvilli of the secretory cell to the exterior differ in the three gland types. The significance of the numbers of cells comprising gland units is related to the role of cells in regenerating the cuticular components of the glands at moulting in nymphs.     

Secretory Vesicle Pools and Rate and Kinetics of Single Vesicle Exocytosis in Neurosecretory Cells

Secretory vesicles are localized in specific compartments within neurosecretory cells. Morphometric, cytochemical and electrophysiological techniques have allowed the definition of secretory vesicle compartments. These are different pools in which vesicles are in various states of releasability. The transit of vesicles between compartments is not random, but an event controlled and regulated by Ca²⁺ and the cortical F-actin network. Cortical F-actin disassembly, a Ca²⁺-dependent event, controls the transit of secretory vesicles from the reserve compartment to the release-ready vesicle pool. Furthermore, the recent development of new technical approaches (patch-clamp membrane capacitance, electrochemical detection of amines with carbon-fibre microelectrodes) has now permitted us to understand the kinetics of single vesicle exocytosis.     

Elektronenmikroskopische Untersuchungen über die Sinneskante des terricolen Turbellars Bipalium kewense Moseley (Tricladida)

Zusammenfassung Die Sinneskante des landlebenden Turbellars Bipalium kewense setzt sich aus folgenden Zellformen zusammen: Papillen und distaler Teil der Epithelfalten werden von einem versenkten Epithel bedeckt. Die distale Cytoplasmalage ist besonders dicht und durch ein tubuläres System gekennzeichnet, das offenbar aus dem Golgi-Apparat der Perikaryen hervorgegangen ist. Die Perikaryen sind tief versenkt und enthalten außer dem Kern granuläres ER, Mitochondrien, Golgi-Apparat und vor allem viel Glykogen. Die Epidermis ist zellig organisiert. In ihrem Bereich kommen drei Sekretzell-Formen vor, die ebenfalls weit in die unterlagernde Bindegewebs-Muskel-Lage hineinragen.Die Sinneskante ist durch zwei Receptor-Typen charakterisiert. Multipolare Zellen erreichen mit mehreren Dendriten die Körperoberfläche im Bereich der Papillen und dem äußeren Teil der Furchen des Kopfvorderrandes. Bipolare Zellen finden sich nur tief eingesenkt in den Sinnesgruben. In ihrem Bereich kommen keine Stützzellen vor. Außer durch die Gestalt sind beide Receptoren durch die Struktur ihrer Cilien unterschieden. Wabendesmosomen sind beiden gemeinsam.

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Electron microscopic observations on the sensory border of the land-dwelling turbellarian Bipalium kewense moseley (tricladida)

Summary The sensory border of the land-dwelling turbellarian Bipalium kewense consists of the following cell types: Papillae and distal part of the epithelial folds are covered by epithelial cells, the perikarya of which lie deeply below the basal lamina. Their distal cytoplasm is particularly dense and characterized by a tubular system, which obviously originates from the Golgi apparatus. The perikarya contain beside the nucleus mitochondria, rough endoplasmic reticulum, Golgi apparatus and above all numerous glycogen particles. Within this cellular epidermis three types of secretory cells occur which also penetrate deeply into the underlying layer of connective and muscle tissues.The sensory border is marked by two types of receptor cells. In the area of the papillae and in the outer part of the furrows of the anterior rim of the head, multipolar cells extend several dendrites to the body surface. Bipolar cells have been observed only in the depth of the sensory grooves. In their surroundings no supportive cells occur. Except for their shape the sensory cells differ in the structure of their cilia. Both cell types are interconnected with neighbouring cells by comb-desmosomes.

Herrn Prof. Dr. Drs. h. c. W. Bargmann danke ich für die Überlassung eines Arbeitsplatzes im Anatomischen Institut der Universität Kiel.     

Fine-grained secretory cells in the intestine of the lancelet,Branchiostoma (Amphioxus) lanceolatum,studied by light microscopy

Summary In the posterior part of the mid-gut epithelium in the lancelet, Branchiostoma lanceolatum, fine-grained cells occur. The appearance of the these cells is conspicuous with a basal and an apical swelling with small secretory granules. At the end of the secretion cycle the granular content is released into the gut lumen. The secretion product seems to consist of proteins and probably has an enzyme function. The restricted localization, the fine granulation, and the characteristic shape are features, that make these cells distinguishable from other secretory cells in the lancelet intestinal epithelium. A possible endocrine capacity of the cells is discussed.This work was supported by grant 2124-23 from the Swedish Natural Science Research Council and by grants from the Faculty of Science, University of Stockholm, Sweden.